Determining the physiological status of individual bacterial cells

Abstract

Publisher Summary This chapter determines the physiological status of individual bacterial cells. are used to estimate the physiological status of individual cells in marine and estuarine water samples. (1)The Vital Stain and Probe (VSP) method is an epifluorescent direct microscopy counting technique that combines the general DNA stain DAPI, the vital stain propidium iodide (PI), and 16S rRNA targeted oligonucleotide probes to identify bacterial cells in situ . (2) Nucleoid staining technique in which the nucleoid is observed with epifluorescence microscopy when it is stained with a DNA fluorochrome such as DAPI. (3) Microautoradiography is a technique that detects the cellular localization of a radiolabeled substance ill situ. This method combines microautoradiography with 16S ribosomal RNA-targeted oligonucleotide probes, which allows simultaneous in situ identification and determination of substrate uptake patterns of individual cells. (4) Live/dead baclight tm bacterial viability kit provides a two-color fluorescent assay of bacterial viability. The kit utilizes mixtures of two nucleic acid stains: SYTO 9 and Propidium Iodide (PI). It is a membrane-impermeant stain that can passively diffuse through a cell wall and act as an indicator of loss in membrane integrity and thus as an indicator of cell viability. (5) Direct viable counting [DVC]: Kogure method utilizes the antibiotic nalidixic acid to interfere with normal cell division. In the presence of this antibiotic, sensitive cells grow but do not divide and can therefore be easily identified microscopically as elongated or enlarged cells.